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A PAS domain binds asparagine in the chemotaxis receptor McpB in Bacillus subtilis.

ABSTRACT: During chemotaxis toward asparagine by Bacillus subtilis, the ligand is thought to bind to the chemoreceptor McpB on the exterior of the cell and induce a conformational change. This change affects the degree of phosphorylation of the CheA kinase bound to the cytoplasmic region of the receptor. Until recently, the sensing domains of the B. subtilis receptors were thought to be structurally similar to the well studied Escherichia coli four-helical bundle. However, sequence analysis has shown the sensing domains of receptors from these two organisms to be vastly different. Homology modeling of the sensing domain of the B. subtilis asparagine receptor McpB revealed two tandem PAS domains. McpB mutants having alanine substitutions in key arginine and tyrosine residues of the upper PAS domain but not in any residues of the lower PAS domain exhibited a chemotactic defect in both swarm plates and capillary assays. Thus, binding does not appear to occur across any dimeric surface but within a monomer. A modified capillary assay designed to determine the concentration of attractant where chemotaxis is most sensitive showed that when Arg-111, Tyr-121, or Tyr-133 is mutated to an alanine, much more asparagine is required to obtain an active chemoreceptor. Isothermal titration calorimetry experiments on the purified sensing domain showed a K(D) to asparagine of 14 mum, with the three mutations leading to less efficient binding. Taken together, these results reveal not only a novel chemoreceptor sensing domain architecture but also, possibly, a different mechanism for chemoreceptor activation.

SUBMITTER: Glekas GD

PROVIDER: S-EPMC2804345 | biostudies-literature | 2010 Jan

REPOSITORIES: biostudies-literature

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A PAS domain binds asparagine in the chemotaxis receptor McpB in Bacillus subtilis.

Glekas George D GD Foster Richard M RM Cates Joseph R JR Estrella Jeffrey A JA Wawrzyniak Michael J MJ Rao Christopher V CV Ordal George W GW

The Journal of biological chemistry 20091028 3

During chemotaxis toward asparagine by Bacillus subtilis, the ligand is thought to bind to the chemoreceptor McpB on the exterior of the cell and induce a conformational change. This change affects the degree of phosphorylation of the CheA kinase bound to the cytoplasmic region of the receptor. Until recently, the sensing domains of the B. subtilis receptors were thought to be structurally similar to the well studied Escherichia coli four-helical bundle. However, sequence analysis has shown the ...[more]

PMID: 19864420

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Project description:Motile bacteria sense chemical gradients using chemoreceptors, which consist of distinct sensing and signaling domains. The general model is that the sensing domain binds the chemical and the signaling domain induces the tactic response. Here, we investigated the unconventional sensing mechanism for ethanol taxis in Bacillus subtilis Ethanol and other short-chain alcohols are attractants for B. subtilis Two chemoreceptors, McpB and HemAT, sense these alcohols. In the case of McpB, the signaling domain directly binds ethanol. We were further able to identify a single amino acid residue, Ala431, on the cytoplasmic signaling domain of McpB that, when mutated to serine, reduces taxis to alcohols. Molecular dynamics simulations suggest that the conversion of Ala431 to serine increases coiled-coil packing within the signaling domain, thereby reducing the ability of ethanol to bind between the helices of the signaling domain. In the case of HemAT, the myoglobin-like sensing domain binds ethanol, likely between the helices encapsulating the heme group. Aside from being sensed by an unconventional mechanism, ethanol also differs from many other chemoattractants because it is not metabolized by B. subtilis and is toxic. We propose that B. subtilis uses ethanol and other short-chain alcohols to locate prey, namely, alcohol-producing microorganisms.IMPORTANCE Ethanol is a chemoattractant for Bacillus subtilis even though it is not metabolized and inhibits growth. B. subtilis likely uses ethanol to find ethanol-fermenting microorganisms to utilize as prey. Two chemoreceptors sense ethanol: HemAT and McpB. HemAT's myoglobin-like sensing domain directly binds ethanol, but the heme group is not involved. McpB is a transmembrane receptor consisting of an extracellular sensing domain and a cytoplasmic signaling domain. While most attractants bind the extracellular sensing domain, we found that ethanol directly binds between intermonomer helices of the cytoplasmic signaling domain of McpB, using a mechanism akin to those identified in many mammalian ethanol-binding proteins. Our results indicate that the sensory repertoire of chemoreceptors extends beyond the sensing domain and can directly involve the signaling domain.

Project description:Colonization of plant roots by Bacillus subtilis is mutually beneficial to plants and bacteria. Plants can secrete up to 30% of their fixed carbon via root exudates, thereby feeding the bacteria, and in return the associated B. subtilis bacteria provide the plant with many growth-promoting traits. Formation of a biofilm on the root by matrix-producing B. subtilis is a well-established requirement for long-term colonization. However, we observed that cells start forming a biofilm only several hours after motile cells first settle on the plant. We also found that intact chemotaxis machinery is required for early root colonization by B. subtilis and for plant protection. Arabidopsis thaliana root exudates attract B. subtilis in vitro, an activity mediated by the two characterized chemoreceptors, McpB and McpC, as well as by the orphan receptor TlpC. Nonetheless, bacteria lacking these chemoreceptors are still able to colonize the root, suggesting that other chemoreceptors might also play a role in this process. These observations suggest that A. thaliana actively recruits B. subtilis through root-secreted molecules, and our results stress the important roles of B. subtilis chemoreceptors for efficient colonization of plants in natural environments. These results demonstrate a remarkable strategy adapted by beneficial rhizobacteria to utilize carbon-rich root exudates, which may facilitate rhizobacterial colonization and a mutualistic association with the host. Bacillus subtilis is a plant growth-promoting rhizobacterium that establishes robust interactions with roots. Many studies have now demonstrated that biofilm formation is required for long-term colonization. However, we observed that motile B. subtilis mediates the first contact with the roots. These cells differentiate into biofilm-producing cells only several hours after the bacteria first contact the root. Our study reveals that intact chemotaxis machinery is required for the bacteria to reach the root. Many, if not all, of the B. subtilis 10 chemoreceptors are involved in the interaction with the plant. These observations stress the importance of root-bacterium interactions in the B. subtilis lifestyle.

Dataset Information

A PAS domain binds asparagine in the chemotaxis receptor McpB in Bacillus subtilis.

Publications

A PAS domain binds asparagine in the chemotaxis receptor McpB in Bacillus subtilis.

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